Product description

Ribonuclease A (Ribonuclease A, commonly abbreviated as RNase A) is a single-chain polypeptide containing four disulfide bonds, with a molecular weight of approximately 13.7 kDa (based on amino acid sequence). As an endoribonuclease, it specifically degrades cytidine (C) or uridine (U) residues in single-stranded RNA. Specifically, it recognizes and cleaves the phosphodiester bond formed between the 5'-ribose of one nucleotide and the 3'-phosphate group of an adjacent pyrimidine nucleotide, resulting in hydrolysis of the 2',3'-cyclic phosphate to yield the corresponding 3'-nucleotide monophosphate (for example, pG-pG-pC-pA-pG is cleaved by RNase A to produce pG-pG-pCp and A-pG). RNase A exhibits the highest activity against single-stranded RNA. The recommended working concentration is 1-100 μg/mL, and it is compatible with various reaction systems.

Under low salt concentrations (0-100 mM NaCl), it can cleave single-stranded RNA, double-stranded RNA, and the RNA strand in RNA-DNA hybrids. However, under high salt concentrations (≥0.3 M), RNase A specifically cleaves only single-stranded RNA.

The most common application of Ribonuclease A (RNase A) is the removal of RNA during plasmid DNA or genomic DNA preparation. The presence or absence of DNase activity is one of the critical contaminants to consider in this preparation process, and DNase activity can be inactivated by traditional methods such as boiling in a water bath. Additionally, this product can be used in molecular biology applications such as RNase protection assays and RNA sequencing.

Product Properties

Storage

This product should be stored at -25°C to -15°C for 3 years. Transport on ice.

Instructions

[Note]: This is one of the common methods for preparing RNase A stock solution. Alternatively, stock solutions can be prepared using laboratory-established methods or as referenced from literature (e.g., directly dissolving in 10 mM Tris-Cl, pH 7.5, or Tris-NaCl solution).

1) Prepare a 10 mg/mL RNase A stock solution using 10 mM sodium acetate (pH 5.2);

2) Heat at 100°C for 15 min;

3) Cool to room temperature, then add 1/10 volume of 1 M Tris-HCl (pH 7.4) to adjust the pH to 7.4 (e.g., add 500 μL of 1 M Tris-HCl, pH 7.4, to 5 mL of 10 mg/mL RNase stock solution);

4) Aliquot and store at -25°C to -15°C. The solution remains stable for up to 3 years.

[Note]: Boiling RNase A solution under neutral pH may result in precipitation of RNase. When boiled at lower pH, precipitation, if observed, may be due to the presence of protein impurities. If precipitation occurs after boiling, remove impurities by high-speed centrifugation (13,000 rpm), then aliquot and store at -25°C to -15°C.

Notes

1. This product has been treated during production to remove DNase contamination. For routine plasmid DNA or genomic DNA extraction applications (where strict quantification of DNA levels is not required), high-temperature boiling is not necessary; the stock solution can be prepared directly. For experiments requiring strict control of residual DNase activity, high-temperature boiling is recommended, or use DNase-free, protease-free RNase A solution (10 mg/mL) (Catalog Number: 10405ES03).

2. RNase A strongly adsorbs to glassware; it is recommended to store the solution in plastic centrifuge tubes.

3. Researchers simultaneously working with intact RNA should be extremely cautious to prevent RNase A from interfering with experimental results.

4. For your safety and health, please wear a lab coat and disposable gloves when handling this product.

5. This product is intended for research use only.